Calcium glycerophosphate for treating and preventing respiratory diseases or conditions

ABSTRACT

Calcium glycerophosphate is found to be effective in treating and preventing a disease, disorder and/or condition of the respiratory system. The disease, disorder and/or condition is related to an obstructive or a restrictive condition of the respiratory airway. The disease, disorder and/or condition can be a respiratory airway inflammatory disease, a respiratory airway stenosis or a nasal cavity inflammatory disease, such as an asthma, a chronic obstructive pulmonary disease (COPD), an emphysema, a reactive airway disease (RADS), rhinitis, bronchitis, bronchiolitis, congestion, sinusitis, tonsillitis, or laryngitis, post-nasal drip (PND) and a related complication thereof, inflamed degranulating and non-degranulating mast cell activity, any irritation occasioning mucus secretion from goblet cells breathing difficulty, restriction, obstruction; airways constriction or closure or mucus interference with air passage; sleep apnea, snoring, inflammatory or non-inflammatory responses to an airborne or non-airborne allergen or irritant; nasal or non-nasal airway inflammation or irritation caused by a problem in any area of the body; and a physical damage to the respiratory system. Methods, compositions and devices are described for using calcium glycerophosphate to treat and prevent the disease, disorder and/or condition of the respiratory system.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a continuation-in-part patent application ofInternational Patent Application No. PCT/US08/74589, filed on Aug. 28,2008, which was published as WO 2009/029705 on Mar. 5, 2009, titled“Calcium Glycerophosphate for Treating and Preventing RespiratoryDiseases or Conditions,” which claims the benefit of U.S. ProvisionalPatent Application No. 60/968,938, filed Aug. 30, 2007, the disclosureof which is herein incorporated by reference in its entirety. Thisapplication also claims the benefit of U.S. Provisional PatentApplication No. 61/285,260, filed Dec. 10, 2009, the disclosure of whichis herein incorporated by reference in its entirety.

BACKGROUND OF THE INVENTION

Diseases, disorders and/or conditions of the respiratory system occurcommonly in both affluent countries and developing countries. Theyaccount for a significant proportion of all days of sickness relatedabsence from work. The morbidity related to respiratory diseases,disorders and conditions has not decreased.

Therefore, there is a need to develop a relatively inexpensive means fortreating and preventing diseases, disorders and/or conditions of therespiratory system. Preferably, such a means is non-toxic, non-hazardousand without significant side effects.

BRIEF SUMMARY OF THE INVENTION

It is now discovered that calcium glycerophosphate is effective intreating and preventing a disease, disorder and/or condition of therespiratory system.

In one general aspect, the present invention relates to a method oftreating or preventing a disease, disorder and/or condition of therespiratory system in a subject. The method comprises administering tothe respiratory system of the subject an effective amount of calciumglycerophosphate in a composition formulated for oral or nasaladministration.

In another general aspect, the present invention relates to acomposition for treating or preventing a disease, disorder and/orcondition of the respiratory system in a subject. The compositioncomprises an effective amount of calcium glycerophosphate and isformulated for oral or nasal administration to the respiratory system ofthe subject by a nasal drop, a nasal spray, a gel, a nasal lavage, aquick-dissolving tablet, an inhaled powder, an oral inhalation solutionor suspension, a syrup, a mechanized intermittent fluid pulser (such asWater-Pik®), an inhaler, a respirator, a transpirator, an atomizer, avaporizer, a nebulizer, an air mask, an insufflator, a means for directphysical or mechanical application (such as a cotton swab), etc.

In yet another general aspect, the present invention relates to a devicefor treating or preventing a disease, disorder and/or condition of therespiratory system in a subject. The device comprises an effectiveamount of calcium glycerophosphate and a means for administering theeffective amount of calcium glycerophosphate to the respiratory systemof the subject.

Another general aspect of embodiments of the invention relates to amethod of reducing, interdicting and/or repairing irritation to therespiratory system of a subject by an agent administered to therespiratory system of the subject, the method comprising:

-   -   1) administering the agent to the respiratory system of the        subject; and    -   2) administering an effective amount of calcium glycerophosphate        to the respiratory system of the subject,        wherein the effective amount of calcium glycerophosphate        reduces, interdicts and/or repairs irritation to the respiratory        system of the subject by the agent.

Other aspects, features and advantages of the invention will be apparentfrom the following disclosure, including the detailed description of theinvention and its preferred embodiments and the appended claims.

BRIEF DESCRIPTION OF THE DRAWINGS

The foregoing summary, as well as the following detailed description ofthe invention, will be better understood when read in conjunction withthe appended drawings. For the purpose of illustrating the invention,there are shown in the drawings embodiments of the invention. It shouldbe understood, however, that the invention is not limited to the precisearrangements and instrumentalities shown.

In the drawing:

FIG. 1 summarizes the study design for characterizing the effects of CGPon rhinitis in ragweed sensitized Beagle dogs;

FIG. 2 illustrates percentage of nasal cavity volume relative to that ofbaseline (time 0) measured before and after intranasal ragweedchallenge: nasal congestion was measured three days after firstinhalation and IN treatment with vehicle or CGP (1.5 mg inhale+30 mgIN); dogs served as their own control and vehicle experiment wasperformed about 5 weeks prior to compound treatment; data are expressedas mean±sem (n=5); nasal congestion was significantly attenuatedcompared to vehicle treatment determined by Two Way Anova (p=<0.0001)followed by Bonferroni posttest at individual time points (** p<0.01);

FIG. 3 shows the area under the curve (AUC) of the change in nasalcavity volume between 0 and 90 min as shown in FIG. 2 expressed asmean±sem (A, n=5) and as scatter graph for individual dogs (B), the AUCwas significantly increased after CGP treatment compared to vehiclecontrol determined by Paired t-test (p=0.0092) indicating an attenuationof the nasal congestion induced by ragweed instillation;

FIG. 4 shows histamine levels in nasal lavage fluid measured before andafter intranasal ragweed challenge done three days after first treatmentwith vehicle or compound: data are expressed as mean±sem (n=5); T₀indicates baseline sample collected the day before the initiation ofeither vehicle or calcium glycerophosphate treatment; dogs served astheir own control and vehicle treatment was compared to compoundtreatment; no statistical significance was reached by an analysis underOne Way Anova followed by Dunnett's posttest;

FIG. 5 shows the AUC calculated for the histamine levels between 0 and60 min in FIG. 4 for vehicle and compound treatment expressed asmean±sem (A, n=5) and individual data points in a scatter graph (B);histamine levels decreased in two of the 5 dogs, no statisticalsignificance was reached;

FIG. 6 shows levels of leukotriene C₄/D₄/E₄ in nasal lavage fluidmeasured before and after intranasal ragweed challenge done three daysafter first treatment with vehicle or compound; data are expressed asmean±sem (n=5); T₀ indicates baseline sample collected the day beforethe initiation of either vehicle or calcium glycerophosphate treatment;dogs served as their own control and vehicle treatment was compared tocompound treatment; no statistical significance was reached by ananalysis under One Way Anova followed by Dunnett's posttest;

FIG. 7 shows the AUC calculated for the levels of leukotriene C₄/D₄/E₄between 0 and 60 min in FIG. 6 for vehicle and compound treatment shownas mean±sem (A, n=5) and individual data points in a scatter graph (B);leukotriene levels decreased in three dogs and increased in two dogs andno statistical significance was reached;

FIG. 8 shows prostaglandin D₂ levels in nasal lavage fluid measuredbefore and after intranasal ragweed challenge done three days afterfirst treatment with vehicle or compound; data are expressed as mean±sem(n=5); T₀ indicates baseline sample collected the day before theinitiation of either vehicle or calcium glycerophosphate treatment; dogsserved as their own control and vehicle treatment was compared tocompound treatment; no statistical significance was reached by ananalysis under One Way Anova followed by Dunnett's posttest;

FIG. 9 shows the AUC calculated for the prostaglandin D₂ levels between0 and 60 min in FIG. 8 for vehicle and compound treatment shown asmean±sem (A, n=5) and individual data points in a scatter graph (B);prostaglandin D₂ levels decreased in three dogs and increased in twodogs and no statistical significance was reached;

FIG. 10 shows prostaglandin E₂ levels in nasal lavage fluid measuredbefore and after intranasal ragweed challenge done three days afterfirst treatment with vehicle or compound; data are expressed as mean±sem(n=5); T₀ indicates baseline sample collected the day before theinitiation of either vehicle or calcium glycerophosphate treatment; dogsserved as their own control and vehicle treatment was compared tocompound treatment; no statistical significance was reached by ananalysis under One Way Anova followed by Dunnett's posttest;

FIG. 11 shows that after the data in FIG. 10 were normalized to takeinto account of the differences in the initial baseline levels (T₀) ofprostaglandin E₂ prior to treatment (e.g. baseline=100%), prostaglandinE₂ levels were significantly attenuated compared to vehicle treatmentdetermined by Two Way Anova (p=<0.009) followed by Bonferroni posttestat individual time points (no significance at individual time points);

FIG. 12 shows the total numbers of cells in nasal lavage (includingepithelial cells) collected before and after repeated oral treatmentwith vehicle and calcium glycerophosphate; nasal lavages were performedprior to first treatment and on Days 1 and 2 after nasal RW challenge on4^(th) day of treatment;

FIG. 13 shows epithelial cells in nasal lavage expressed as percentageof total cells (A) and as number of cells (B) collected before and afterrepeated oral treatment with vehicle and calcium glycerophosphate; nasallavages were performed prior to first treatment and on Days 1 and 2after nasal RW challenge on 4^(th) day of treatment;

FIG. 14 shows neutrophils in nasal lavage expressed as percentage oftotal cells (A) and as number of cells (B) collected before and afterrepeated oral treatment with vehicle and calcium glycerophosphate; nasallavages were performed prior to first treatment and on Days 1 and 2after nasal RW challenge on 4^(th) day of treatment;

FIG. 15 shows eosinophils in nasal lavage expressed as percentage oftotal cells (A) and as number of cells (B) collected before and afterrepeated oral treatment with vehicle and calcium glycerophosphate; nasallavages were performed prior to first treatment and on Days 1 and 2after nasal RW challenge on 4^(th) day of treatment;

FIG. 16 shows macrophages in nasal lavage expressed as percentage oftotal cells (A) and as number of cells (B) collected before and afterrepeated oral treatment with vehicle and calcium glycerophosphate; nasallavages were performed prior to first treatment and on Days 1 and 2after nasal RW challenge on 4^(th) day of treatment; and

FIG. 17 shows lymphocytes in nasal lavage expressed as percentage oftotal cells (A) and as number of cells (B) collected before and afterrepeated oral treatment with vehicle and calcium glycerophosphate; nasallavages were performed prior to first treatment and on Days 1 and 2after nasal RW challenge on 4^(th) day of treatment.

DETAILED DESCRIPTION OF THE INVENTION

Calcium glycerophosphate has already been shown to behave as ananti-inflammatory substance on epidermal and epithelial cells and as awound healer on epidermal cells as well as in the gums and mucosal softtissue elsewhere in the body, e.g., vaginal. Investigation has beenexpanded to its use on the nasal mucosa and other parts of therespiratory system.

Unless defined otherwise, all technical and scientific terms used hereinhave the same meaning as commonly understood to one of ordinary skill inthe art to which this invention pertains. Otherwise, certain terms usedherein have the meanings as set in the specification. All patents,published patent applications and publications cited herein areincorporated by reference as if set forth fully herein. It must be notedthat as used herein and in the appended claims, the singular forms “a,”“an,” and “the” include plural reference unless the context clearlydictates otherwise.

As used herein, the term “subject” refers to a mammal, who has been theobject of treatment, observation or experiment. Examples of a subjectcan be a human, a livestock animal (beef and dairy cattle, sheep,poultry, etc.), or a companion animal (dog, cat, horse, etc).

As used herein, the term “respiratory system” refers to all parts of theairway, i.e., the passageway for air during respiration, from the noseto the pulmonary alveoli. The respiratory system includes organs thatare involved in breathing, such as the nose, throat, larynx, trachea,bronchi, and lungs.

As used herein, the term a “disease, disorder and/or condition of a/therespiratory system” refers to any disease, disorder and/or conditionthat is related to an obstructive or restrictive condition of arespiratory system. An obstructive condition of a respiratory systemincludes any condition which impedes the rate of air flow into and outof the lung. A restrictive condition of a respiratory system includesany condition which causes a reduction in the functional volume of thelung. The obstruction or restriction of the airway may cause symptomssuch as wheezing, shortness of breath, difficulty breathing, chesttightness, and coughing. The disease, disorder and/or condition of therespiratory system can be, for example, an airway inflammatory disease,an airway stenosis, or a nasal cavity inflammatory disease.

Examples of the disease, disorder and/or condition of the respiratorysystem include, but are not limited to, an asthma; a chronic obstructivepulmonary disease (COPD); an emphysema, a reactive airway disease(RADS); rhinitis; bronchitis; bronchiolitis; congestion; sinusitis;tonsillitis; laryngitis; post-nasal drip (PND) and any and allcomplications dependent on same; inflamed degranulating andnon-degranulating mast cell activity; any irritation occasioning mucussecretion from goblet cells or elsewhere, resulting in breathingdifficulty, restriction and/or obstruction; airway constriction orclosure or mucus interference with air passage; sleep apnea; snoring;inflammatory or non-inflammatory responses to any airborne or otherallergen or irritant; nasal or other airway inflammation or irritationcaused by any other body area problem; physical damage to therespiratory system such as nosebleed, surgery healing, traumatic injury;any respiratory disease, disorder and/or condition caused by an airborneor seasonal allergen or irritant, any swelling of tissue occasioned byany of the above, etc.

As used herein, the term “asthma” refers to a chronic condition, whichin most cases is characterized by reversible airway obstructions and/orconstrictions. The airway becomes inflamed and is lined with excessiveamounts of mucus, often in response to one or more triggers for asthma.The triggers for asthma include, but are not limited to, anenvironmental stimulant, such as an allergen (ragweed, house dust,animal hair, pollen, etc.), cold air, warm air, moist air, change intemperature or humidity, upper respiratory infections, exercise,exertion, physical or emotional stress, smoke, viral illnesses such asthose caused by common cold. The term “asthma” includes those caused byany cause of asthma whose primary effect is cellular inflammation and/orirritation, whether involving mast cells or not, degranulation or not,mucus exudation or not, whether exacerbant is identified or not, orwhether the cause is airborne or not. The term ‘asthma’ is to be thewidest-encompassing and is to include breathing difficulty of alldegrees from the barely perceptible to acute.

Examples of asthma include, but are not limited to bronchial asthma,infantile asthma, allergic asthma, atopic asthma, steroid refractoryasthma, non-allergic asthma, endogenous asthma, exogenous asthma,aspirin asthma, cardiac asthma, exercise-induced asthma, infectiousasthma, any asthma triggered by airway restriction or constriction.

As used herein, the term “chronic obstructive pulmonary disease” or“COPD”, also known as chronic obstructive airway disease (COAD), refersto a progressive respiratory disease characterized by limitation ofairflow in the airway that is not fully reversible. COPD often involvespermanent or temporary narrowing of small bronchi, in which forcedexpiratory flow is slowed. Examples of COPD include chronic bronchitis,emphysema and a range of other disorders to which no etiologic or othermore specific term can be applied. COPD is most often due to tobaccosmoking but can be due to other airborne irritants, such as coal dust,asbestos or solvents, as well as preserved meats containing nitrites.

As used herein, the term “reactive airway disease (RAD)” refers to anasthma-like syndrome developed after a single exposure to high levels ofa trigger, such as irritating vapor, fume, or smoke. In a particularembodiment of the present invention, the term RAD includes anasthma-like syndrome in infants that may later be confirmed to be asthmawhen they become old enough to participate in diagnostic tests.

As used herein, the term “rhinitis” refers to any disease, disorderand/or condition caused by inflammation of the nasal mucous membrane.Examples of rhinitis include, but are not limited to, allergic rhinitis,pollinosis, acute rhinitis, chronic rhinitis, hypertrophic rhinitis,deflected septum and the like. Symptoms of rhinitis include, but are notlimited to, a runny nose, nasal congestion and post-nasal drip.According to recent studies completed in the United States, more thanfifty million Americans are current sufferers of rhinitis. Rhinitis hasbeen found to adversely affect more than just the nose, throat, andeyes. It has been associated with sleeping problems, problems with theears, and has even been linked to learning problems. Causes that maybring about the presence of rhinitis include food reactions, anatomicdefects, immunodeficiency diseases, ciliary dyskinesia, environmentaltriggers, emotional triggers, occupational triggers, hormonal triggers,etc.

As used herein, the term “calcium glycerophosphate” or “CGP,” also knownas “glycerophosphate calcium,” refers to a chemical compound having amolecular formula of C₃H₇CaO₆P in its anhydrous form. “CGP” can alsoexist as a hydrate, including the monohydrate and the dihydrate.Examples of calcium glycerophosphate include, but are not limited to,any one, or any combination of two or more of the three isomers of CGP,namely β-glycerophosphoric acid calcium salt ((HOCH₂)₂CHOPO₃Ca) and D(+)and L(−)-α-glycerophosphoric acid calcium salt (HOCH₂CH(OH)CH₂OPO₃Ca).

Calcium glycerophosphate can be synthesized using methods known in theart. Calcium glycerophosphate can also be obtained from variouscommercial sources. The commercially available CGP preparations include,but are not limited to, those available from AkPharma Inc.(Pleasantville, N.J. 08232), Astha Laboratories Pvt, Ltd, (B-4,Industrial Estate, Sanathnagar, Hyderabad-18, India), and Seppic Inc.(30 Two Bridges Road, Fairfield, N.J. 07004).

As used herein the term “treatment”, “treat” or “therapy” refers to theprevention of deterioration of a disease, disorder or condition when apatient contracts such a disease, disorder or condition, preferably, atleast maintenance of the status quo, and more preferably, alleviation,still more preferably, resolution of the disease, disorder or condition.

As used herein the term “prophylaxis”, “prevent” or “prevention” refersto, when referring to a disease, disorder or condition, a type oftreatment conducted before such a disease, disorder or condition occurssuch that the disease, disorder or condition will not occur, will bedelayed to occur, or will occur but will deteriorate to a less degree.

As used herein, the term “treat” or “prevent” in the broadest sense,with respect to a disease, disorder or condition, refers to any medicalact thereto, and include any act for diagnosis, therapy, prevention,prognosis and the like.

When used for treating or preventing a disease, disorder and/orcondition of the respiratory system, calcium glycerophosphate can beused as a reliever which is used during an episode or an attack of thedisease, disorder and/or condition, such as an episode of an asthma, foralleviation of the episode or attack. Calcium glycerophosphate can alsobe used as a controller which is used for long-term control to preventthe occurrence of the episode or attack. Controlling or preventing anattack is substantially the therapy of a disease, disorder and/orcondition of the respiratory system per se, because it is equallyimportant to control and prevent an attack as to relieve or alleviatethe attack. Those skilled in the art will be able to use an appropriatedosage of calcium glycerophosphate for either therapy or prevention of adisease, disorder and/or condition of the respiratory system.

The term “effective amount” as used herein means that amount of activecompound or pharmaceutical agent that elicits the biological ormedicinal response in a tissue system, animal or human that is beingsought by a researcher, veterinarian, medical doctor or other clinician,which includes alleviation of the symptoms of the disease or disorderbeing treated. The administration of an effective amount of calciumglycerophosphate to a subject results in a clinically observablebeneficial effect. The clinically observable beneficial effect can be asituation in which an observable disease, disorder and/or condition ofthe respiratory system is prevented from further development oraggravation or will develop to a lesser degree, than withoutadministration of the composition of the present invention. Theclinically observable beneficial effect can also be a situation in whicha disease, disorder and/or condition of the respiratory system isprevented from occurring or subsequently occurs to a lesser degree thanwithout administration of the composition of the present invention, whenthe composition is administered to a subject before the disease,disorder and/or condition of the respiratory system is observable. Inone embodiment of the invention, an effective amount of calciumglycerophosphate alleviates or improves a disease, disorder and/orcondition of the respiratory system in a subject to a degree that isabout any of 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, or 90% of thatwhich would have been had the subject not received an effective amountof calcium glycerophosphate.

Methods are known in the art for determining therapeutically andprophylactically effective doses of calcium glycerophosphate accordingto embodiments of the present invention. A useful assay for confirmingan effective amount (e.g., a therapeutically effective amount) for apredetermined application is to measure the degree of recovery from atarget disease. An amount actually administered depends on an individualto be treated. The amount is preferably optimized so as to obtain adesired effect without significant side effects. The determination of aprophylactically or therapeutically effective dose is within the abilityof those skilled in the art. A prophylactically or therapeuticallyeffective dose of any compound can be estimated using either a cellculture assay or any appropriate animal model. The animal model is usedto achieve a desired concentration range and an administration route.Thereafter, such information can be used to determine a dose and routeuseful for administration into humans.

The therapeutic effect and toxicity of a compound may be determined bystandard pharmaceutical procedures in cell cultures or experimentalanimals (e.g., ED₅₀, a dose therapeutically effective for 50% of apopulation; and LD₅₀, a dose lethal to 50% of a population). The doseratio between therapeutic and toxic effects is a therapeutic index, andit can be expressed as the ratio of LD₅₀/ED₅₀. Pharmaceuticalcompositions which exhibit high therapeutic indices are preferable. Thedata obtained from cell culture assays and animal studies can be usedfor formulating a dosage range for use in humans. The dosage of suchcompounds lies preferably within a range of circulating concentrationsthat include the ED₅₀, with little or no toxicity. Such a dosage mayvary within this range depending upon the dosage form employed, thesusceptibility of a patient, and the route of administration. Guidancefor specific doses and delivery methods is provided in publicationsknown in the art. The exact dose is chosen by an individual physician inview of the condition of a patient to be treated. Doses andadministration are adjusted to provide a sufficient level of the activeportion, or to attain a desired effect.

The effective amount of CGP can be any dosage amount, from micro-dosesto mega-doses. Mega-doses of CGP can be effectively used, because CGP isnon-toxic, non-hazardous and has no known side effects. Micro-doses ofCGP can be effectively used, because both Ca²⁺ and the glycerophosphateanion are signaling molecules that can have a biological effect at verylow levels.

In embodiments of the present invention, the effective amount of CGP isadministered to the subject in a composition containing about 0.05%-15%(w/w), preferably about 0.5%-10% (w/w); most preferably about 1%-5%(w/w) of CGP. It has been discovered that at higher levels tested, e.g.,about 7.5% (w/w) or above, calcium glycerophosphate may help instanching nosebleeds, possibly due to the effect of calcium on bloodclotting. However, the amount of CGP in the composition is not limitedto about 0.05%-15% (w/w).

In one general aspect, an embodiment of the present invention relates toa method of treating or preventing a disease, disorder and/or conditionof a respiratory system in a subject. The method comprises administeringto the respiratory system of the subject an effective amount of calciumglycerophosphate, wherein calcium glycerophosphate is administered tothe respiratory system of the subject in a composition formulated fororal or nasal administration. The composition formulated for oral ornasal administration can be a liquid, solid, gel, syrup, powder, or mistformulation.

Calcium glycerophosphate can be administered to the respiratory systemof the subject by one or more means of oral or nasal administrationdepending on the type of diseases, disorders and/or conditions of therespiratory system. For example, in the case of asthma, COPD and thelike, atomizer type inhalators such as MDI, BDI, or nebulizers and thelike may be used for inhalation. For example, in the case of rhinitis,absorption and inhalation may be used for administration. Examples ofapplicable means of oral or nasal administration include, but are notlimited to, a nasal drop, a nasal spray, a nasal lavage, aquick-dissolving tablet, an inhaled powder, an oral inhalation solutionor suspension, a syrup, a mechanized intermittent fluid pulser (such asWater-Pik®), an inhaler, a respirator, a transpirator, an atomizer, avaporizer, an air mask, an insufflator, a means for direct physical ormechanical application, such as a cotton swab, etc.

Inhalation is conventionally used as a method for administration via thenasal cavity, airway and nasal pathways and the like. In intra-airwayadministration formulations, transairway absorption formulations orpernasal absorption formulations, it is usually preferable to make adrug solution in a mist form or as fine powder (dry powder). Generally,a drug solution formed may be inhaled by means of a nebulizer, thoseprocessed into powder may be inhaled by means of a gas-atomizing type,MDI (metered dose inhaler) or expiration inhalation system, DPI (drypowder inhaler) with the drug loaded therein.

With respect to powder inhalers, there are two types presently used forrapid and deep inhalation, “dry powder inhaler (DPI)” and delayedinhaling type “metered dose inhaler (MDI)”. DPI are further classifiedinto three categories: multi-dose reservoir, such as the productTURBUHALER®, available from AstraZeneca; multi-unit dose, such as theproduct ACCUHALER™/FLOVENT DISKUS™, Advair™, available from GSK; andunit dose, available from many manufacturers.

Inhaler refers to a kit comprising a mouth piece and cartridge (tube),and are usually employed by sealing both termini of the tube withaluminum foil. Prior to use, the tube is equipped with the mouth pieceto pierce the aluminum foil, thereby allowing inspiration of powdereddrug inside.

On the other hand, absorption of a drug solution may be achieved bymeans of a nebulizer or respirator, an artificial respirator. Anebulizer causes a drug aerosol to flow in the air at slow speed, thusmakes it easier for one to absorb the drug.

How often and how long calcium glycerophosphate is administered to asubject depends on the disease, disorder and/or condition of therespiratory system to be treated or prevented, as well as factorsassociated with the subject, e.g., age, weight, health, etc. Calciumglycerophosphate can be administered on a regimen of one to multipletimes per day. Calcium glycerophosphate can be administered to thesubject at intervals during the day, such as upon arising, afterbreakfast, lunch, dinner, and upon retiring. Calcium glycerophosphatecan be administered during an episode or an attack of the disease,disorder and/or condition of the respiratory system to provide a reliefof symptoms, such as wheezing, shortness of breath, difficultybreathing, chest tightness, and coughing. Calcium glycerophosphate canalso be administered to a subject prior to an episode or an attack tocontrol or prevent the episode or attack and the symptoms associatedwith the episode or attack.

Dosages of calcium glycerophosphate are not limited to a particularvalue. The dosage appropriately varies depending on the targeteddisease, condition (extent), age, the presence or absence ofcomplication(s), etc. For example, the dosage is usually, per adult, peradministration, about 100 μg to about 1000 mg, preferably about 500 μgto about 100 mg, and most preferably about 1 mg to about 40 mg ofanhydrous CGP. As used herein, “anhydrous CGP” refers to a CGPpreparation that contains at least about 88% (w/w) of CGP that is freeof residual or acquired moisture. The anhydrous CGP used in embodimentsof the present invention complies with Food Chemicals Codex (FCC)specifications, in which loss on drying (LOD) is not to exceed 12%. Asused herein, “per administration” can be, per inhalation per nostril,per spray, per tablet, etc. In one embodiment of the present invention,the dosage is about 400 mg solution/suspension formulation containingabout 2% by weight of dry CGP per administration. In another embodimentof the present invention, the dosage is about 400 mg solution/suspensionformulation containing about 3.75% by weight of dry CGP peradministration. The dry CGP contains about 95-98% anhydrous CGPbalancing with moisture. However, the dosage of CGP is not limited tothe above ranges and can be any range without causing physicalendangerment.

While not wishing to be bound by theory, calcium glycerophosphate can beused to treat or prevent a disease, disorder and/or condition of therespiratory system at least in part due to the anti-inflammatory effectof CGP. Inflamed airway epithelium results in a disease, disorder and/orcondition of the respiratory system. Various observations suggest that aglycerophosphate salt functions to promote epidermal cell renewal, seefor example, US2004/0037766. The quick repair and replacement ofepidermal cells provide, among other things, enhanced ceramidesynthesis, which hastens repair of the skin's surface and providestighter cell-to-cell adhesion, which may prevent invasion betweenvulnerable cell walls of irritating substances. This is to bedistinguished from the possible function of the calcium ion to modifythe permeability of cell membranes, per se, i.e., the ability of calciumion to decrease membrane porosity at a large concentration. Thereduction, interdiction, suppression or prevention of inflammation ofthe respiratory system provides symptom relief or prevention. Thebeneficial effect of CGP may also be due to, at least in part, itsability to prevent or reduce acid-caused irritation and cytotoxicity inthe upper and lower respiratory tract, and/or its ability to promotehigher ciliary activity, e.g., via regulating the phosphorylation stateof certain ciliary proteins. It is believed that the newly discoveredbeneficial effect of CGP on the respiratory system according toembodiments of the present invention is achieved synergistically betweenthe calcium ion and the glycerophosphate. This synergistic effect isdistinct from the function of the calcium ion or the glycerophosphatealone.

Calcium glycerophosphate is non-toxic, non-hazardous and has no knownside effects. Therefore, methods according to embodiments of the presentinvention are particularly desirable for pediatric patients, elderlypatients, pregnant women, or patients who have frequent need of reliefmedications and/or preventive medications for a disease, disorder and/orcondition of the respiratory system. Oral or nasal administration of thecomposition according to the present invention is non-invasive and canbe repetitively provided.

In particular embodiments, calcium glycerophosphate can be administeredin combination with one or more other relief and/or preventive agentsfor a disease, disorder and/or condition of the respiratory system.Thus, embodiments of the present invention relate to compositionscomprising calcium glycerophosphate and one or more other relief and/orpreventive agents for a disease, disorder and/or condition of therespiratory system, and methods of using the compositions for treatingor preventing a disease, disorder and/or condition of a respiratorysystem in a subject. Calcium glycerophosphate and the other agent can beadministered simultaneously or sequentially, one following the other.The other agents can be administered to the subject via routes ofadministration customarily used for such other drugs. However, it is notnecessary to administer the other relief and/or preventive agent in asubstantial percentage of instances according to embodiments of thepresent invention. Calcium glycerophosphate, as the sole activepharmaceutical ingredient, is effective to treat or prevent a disease,disorder and/or condition of the respiratory system.

Examples of such relief and/or preventive agents include, but are notlimited to, a beta-2 agonist, a long-acting beta-2-agonist (“LABA”), aninhaled corticosteroid, an alpha agonist, a bronchodialator, aglucocorticoid, a leukotriene modifier, a mast cell stabilizer, anantimuscarinic/anticholinergic, a methylxanthine, an antihistamine,omalizumab, methotrexate, and tianeptine, albuterol, cromolyn, or thelike. Examples of LABA include, for example, salmeterol, formoterol,bambuterol, clenbuterol, and indacaterol.

Other embodiments of the present invention relate to compositionscomprising calcium glycerophosphate and one or more analgesics, andmethods of using the compositions for treating or preventing a disease,disorder and/or condition of a respiratory system in a subject. Examplesof the compositions include, but are not limited to, a pharmaceuticalproduct for treating a cold, hayfever, any respiratory disease, disorderand/or condition caused by an airborne or seasonal allergen or irritant,etc., comprising CGP as the nasal cleaner/decongestant (NasoCell™) and acommon over the counter (OTC) analgesic such as ibuprofen,acetominophen, aspirin, naproxen, capsaicin, etc. The amount of CGP inthe composition can be appropriate to supply in a single dosage, whichmay be 2 to 4 sprays of the NasoCell, to provide nasal cleaning and/ordecongestant. The amount of the analgesic in the composition can beeffective to relieve common headaches, sinus aches, eye aches, etc. thatare associated with colds, hay fever, etc.

Nasal administration of the composition according to the presentinvention can provide more rapid relief of the symptoms associated withcold, hayfever, etc. Administration via nasal membrane absorption can bemore quantitatively effective and more chronologically prompt to reachthe bloodstream than the same analgesic ingested that must go throughthe gastric system for subsequent absorption with possible compositionalcompromise by the digestive process. In addition, the hypotonicity ofthe NasoCell allows the composition to adhere more readily to theepithelial nasal cells, thus be absorbed more readily through the cellwalls and into the bloodstream.

The compositions according to the present invention offer the uniquecombination of effective nasal cleaning, nasal clearing,anti-inflammation, anti-swelling, and pain relief, without any of thepsychogenic effects associated with the presently marketed drugs, suchas diphenhydramine, ephedrine, pseudoephedrine, etc., nor any of theundesirable, typical anti-cholinergic side effects at the site orelsewhere in the body. The composition is safe to use liberally evenwhen driving or operating machinery.

In another general aspect, an embodiment of the invention provides acomposition for treating or preventing a disease, disorder and/orcondition of the respiratory system in a subject. The compositioncomprises an effective amount of calcium glycerophosphate, wherein thecomposition is formulated for oral or nasal administration to therespiratory system of the subject by a nasal drop, a nasal spray, anasal lavage, a quick-dissolving tablet, an inhaled powder, an oralinhalation solution or suspension, an inhaler, a respirator, anebulizer, a transpirator, an atomizer, a vaporizer, an air mask, aninsufflator, a means for direct physical or mechanical application, suchas a cotton swab, etc.

Agents for treating or preventing diseases or disorders other thanrespiratory diseases or disorders can also be administered through therespiratory system, such as by nasal administration, because such routeof administration provides faster and more direction absorption of theagent into the blood stream. However, some of the agents may causeirritation or inflammation to the respiratory system. Thus, anothergeneral aspect of embodiments of the invention relates to a method ofreducing, interdicting and/or repairing irritation to the respiratorysystem of a subject by an agent administered to the respiratory systemof the subject. The method comprises

-   -   1) administering the agent to the respiratory system of the        subject; and    -   2) administering an effective amount of calcium glycerophosphate        to the respiratory system of the subject,        wherein the effective amount of calcium glycerophosphate        reduces, interdicts and/or repairs irritation to the respiratory        system of the subject by the agent.

The agent and CGP can be administered simultaneously or sequentially,one following the other. The agent and CGP can be administered in one orseparate formulations for respiratory system administration.

In an embodiment of the present invention, the method of reducing,interdicting and/or repairing irritation to the respiratory system of asubject by an agent administered to the respiratory system of thesubject comprises:

-   1) providing a formulation for respiratory system administration    comprising the agent and an effective amount of calcium    glycerophosphate; and-   2) administering the formulation to the respiratory system of the    subject,    wherein the effective amount of calcium glycerophosphate reduces,    interdicts and/or repairs irritation to the respiratory system of    the subject by the agent.

The formulation can be any formulation administrable to the respiratorysystem of the subject. In an embodiment of the present invention, theformulation is a nasal formulation comprising a therapeutic agent and aneffective amount of CGP. In another embodiment of the present invention,the formulation is for oral inhalation administration that comprises atherapeutic agent and an effective amount of CGP. The effective amountof CGP reduces, interdicts and/or repairs irritation to the respiratorysystem of the subject by the therapeutic agent The formulation isadministered to the respiratory system of the subject by a nasal drop, anasal spray, a nasal lavage, a quick-dissolving tablet, an inhaledpowder, an oral inhalation solution or suspension, a syrup, a mechanizedintermittent fluid pulser, an inhaler, a respirator, a transpirator, anatomizer, a vaporizer, an air mask, a nebulizer, a means for directphysical or mechanical application, or an insufflator.

The composition according to embodiments of the present invention may beproduced using a method similar to methods known in the art, e.g.,conventional mixing, dissolution, rendering to granules, preparation ofa sugar-coated agent, elutriation, emulsification, capsulation,inclusion, or freeze drying. One or more excipients can be added to thecomposition. Excipients which can be used are those that are inactiveagainst calcium glycerophosphate, and as long as the use is recognizedas a pharmaceutical additive, no limitation is made for such excipient.Examples of appropriate excipients include, but are not limited to,monosaccharides such as galactose, mannose, sorbose; disaccharides suchas lactose, sucrose and trehalose and the like; polysaccharides such asstarch, raffinose, dextran and the like; sugar alcohols (includingglycerol, erythritol, arabitol, xylitol, sorbitol, mannitol); glycols(including ethylene glycol, propylene glycol, polyethylene glycol,polypropylene glycol); cellulose-like polymers (including hydroxycellulose, hydroxy propyl cellulose); insoluble additives (crystallinecellulose, chitosan, calcium carbonate, talc, titanium oxide or silica(silicon oxide), and mixtures thereof.

The composition according to embodiments of the present invention can beformulated to have a pH of about 4.5-10, such as about 4.5-6.0, about6.0-7.0, about 7.0-8.0, about 8.0-9.0, about 7.0-9.0, about 7.5-9.0, orabout 8.0-10.0. Note that nasal pH varies considerably, from 4.5-6.5 innormal nasal cells, to as high as 8.3 in rhinitis. However, the pH ofthe composition according to embodiments of the present invention is notlimited to the range of about 4.5-10, or that of the nasal pH.

The composition according to embodiments of the present invention canfurther contain a preservative. Preferably the preservative is foodgrade or pharmaceutical grade. Examples of appropriate preservativesinclude, but are not limited to, methylparaben, ethylparaben,butylparaben, propylparaben, sorbic acid and any other preservative thatis typically used in water-based cosmetics, such as creams and lotionsand some bath products. The preservative is preferably present at anamount that is sufficient to prevent the composition from supporting thegrowth of microbes, such as bacteria, fungi, or yeasts.

The composition according to embodiments of the present invention canalso include an adhesion molecule or material that allows thecomposition to adhere to an airway tissue for an extended period oftime, thus results in an extended release of CGP into the airway.Adherence is accomplished by a number of interactions, physical orchemical, such as electrostatic interaction, hydrogen bonding orhydrophobic interaction. In preferred embodiment, the adhesion moleculeor material extends the contact time of CGP in the nasal cavity. Anysuitable adhesion molecule or material known to a person skilled in theart can be used in a composition according to embodiments of the presentinvention. In one embodiment, the adhesion molecule or material is apolysaccharide. In a preferred embodiment, the adhesion molecule ischitosan, a cationic polysaccharide derived from the shells ofcrustaceans. A versatile transmucosal delivery system based on chitosanis commercially available from West Drug Delivery of Lionville, Pa.19353 (US), and can be used in the present invention.

In a preferred embodiment, the composition of the present is formulatedas a powder, gel, microsphere, or suspension, i.e., liquid comprisingCGP in an amount exceeding the solubility of CGP in the liquid. CGP haslimited solubility in water, i.e., about 1% by weight. Administration ofCGP in a formulation of powder, gel, microsphere, or suspension canresult in a local amount of CGP exceeding its solubility, thusdeposition of insoluble CGP onto the mucous membranes of the respiratorysystem. As the insoluble CGP slowly dissolves into mucous membranes, anextended release of CGP into the cells lining the airway is achievedwithout the need for any additional assisting adhesion substances.Optionally, the powder, gel, microsphere, or suspension formulation caninclude an adhesion molecule, a “sticker”, or material that furtherenhances the CGP's adherence to the airway mucosal surface.

A formulation that provides an extended release of CGP is preferred, forexample, when it is desirable to provide a sustained and steady statelevel of CGP into the respiratory system for an extended time period,such as when the formulation is used for prophylaxis. In one embodiment,a method according to embodiments of the invention comprisesadministering to the respiratory system of the subject a formulationthat provides an extended release of CGP into the airway. In a preferredembodiment, the CGP is released into the airway during a time period of3-8 hours.

In another embodiment, a composition according to embodiment of thepresent invention comprises an absorption enhancer that improvesabsorption of CGP into the airway, e.g., across the mucous membranes ofthe airway. There are a number of ways an absorption enhancer can act.For example, it may alter properties of the mucus layer by opening tightjunctions between the cells, or it may increase membrane fluidity. Anysuitable absorption enhancers known to a person skilled in the art canbe used in a composition according to embodiments of the presentinvention. A formulation that provides an improved absorption of CGP ispreferred, for example, when it is desirable to provide an immediate andhigh level of CGP into the respiratory system, such as when theformulation is used for treatment during an episode or an attack. In oneembodiment, a method according to embodiments of the invention comprisesadministering to the respiratory system of the subject a formulationthat provides an improved absorption of CGP into the airway.

The composition according to embodiments of the present invention can beformulated in various forms that are suitable for oral or nasaladministration in view of the known technologies in the art. Forexample, a form of spray or expiration adapted format includingmicroparticles, such as dry powder, can be used for intra-airwayadministration or transairway absorption. Dry powder may be manufacturedby means of one selected from the group consisting of a bowl mill, abead mill, a jet mill, an ultimizer, a mortar, a stonemill, spray dryingand supercritical fluid. The aerodynamic average particle size of thedry powder may be optimized for administration, e.g., to allow thepowder to float freely in an airstream, settle and adhere to exposedmucosal cell membranes. For airway administration, it is desirable thatthe aerodynamic average particle size of dry powder is typically about0.01 to about 50 μm, preferably, about 0.1 to about 30 μm, stillpreferably about 0.1 to about 10 μm, in diameter. For pulmonaryadministration, in view of the delivery into the alveoli pulmonis, thoseparticles having an aerodynamic average particle size of about 3 μm orless are preferably manufactured, but the present invention is notlimited thereto. The powders can be inhaled or inspired. They can bedispensed via a “puff” container of any sort, including those whichdispense unmetered or metered amounts.

A composition according to an embodiment of the present invention can beassociated with an aerosol system, such as an aerosol spray can. Theaerosol system includes, for example, vessels with propellant includedtherein. The propellant comprises the active ingredient CGP andconventional additives such as lactose. The formulation of thepropellant determines the properties of the output of the aerosolsystem, such as particle distribution, delivery rate, viscosity and thelike. Such aerosol systems can be manufactured using methods known inthe art in view of the present disclosure.

A composition according to an embodiment of the present invention canalso be an aqueous solution/suspension, which is suitable for nasalcavity administration or for inhalation, such as by direct nasal lavage,liquid stream or spray. The solution is atomized to very small particlesof a size range of 1-10 μm. An aerosol finely distributed within anatomized solution can also be inhaled. Detailed information relating toaerosol inhalants is available, for example, from a pharmacopoeia suchas the Japanese Pharmacopoeia, US Pharmacopoeia and the like, which areherein incorporated hereby as references in their entirety. The aqueoussolution/suspension can be sprayed under hand-operated actuation, suchas in a squeezable bottle or plunger, or it can be in a pressurizedcontainer.

A composition according to an embodiment of the present invention canfurther be a gel or cream for nasal application, in which case theingredients would have, in addition to those described above, suitablestabilizing substances to raise the viscosity to desired levels.Examples of such stabilizing substances include, but are not limited to,sodium carboxymethyl cellulose gum (CMC gum), guar gum, xanthan gum,etc. A composition according to an embodiment of the present inventioncan further comprise fatty acids of vegetable source or of animalsource, such as butyric acid.

In a particular embodiment of the present invention, the composition isan aqueous solution/suspension comprising about 1-5% (w/w) CGP; one ormore acceptable moisturizers, such as glycerol, sorbitol and/or thelike; one or more acceptable bacteriostats/mycostats, such as methylparaben, grapefruit seed extract or the like; one or more flavorings oraromatics, such as vanilla, eucalyptus or the like, and purified,sterile water. Such a composition is applied to the respiratory systemby direct nasal lavage or nasal or oral inhalation.

In another particular embodiment of the present invention, thecomposition is an aqueous solution/suspension comprising about 6-10%(w/w) CGP; one or more acceptable moisturizers, such as glycerol,sorbitol and/or the like; one or more acceptablebacteriostats/mycostats, such as methyl paraben, grapefruit seed extractor the like; one or more flavorings or aromatics, such as vanilla,eucalyptus or the like; purified, sterile water; a stabilizing orthickening agent; and an adhesion molecule. Such a composition isapplied to the nasal cavity as a nose drop.

In another aspect, the present invention provides a device for treatingor preventing a disease, disorder and/or condition of the respiratorysystem in a subject. The device comprises an effective amount of calciumglycerophosphate and a means for administering the effective amount ofcalcium glycerophosphate to the respiratory system of a subject. Itshould be understood that such a device may be in any format as long asthe device is for facilitating the administration of calciumglycerophosphate to the respiratory system. The means for administrationto the respiratory system comprises means selected from the groupconsisting of means for administration to the lung, means fortransairway administration, means for transairway absorption and meansfor nasal absorption.

Exemplary means for administration to the airway include, but are notlimited to, an inhaler, a respirator, a transpirator, an atomizer (e.g.,a means to apply a spray/mist from a container such as a squeezed bottleor pressure container), a vaporizer, an air mask, an insufflator, and ameans for direct physical or mechanical application. In particularembodiments, the means for administration to the airway is a metereddose inhaler (MDI), a dry powder inhaler (DPI), a nebulizer, a means forperforming direct nasal lavage, a cotton swab, etc.

Exemplary means for administration to the lung include, but are notlimited to, an inhaler and a bronchoscope, including those bronchoscopesystems commercially available from Olympus Inc.

EXAMPLE 1 Effectiveness of Calcium Glycerophosphate in SensitizedBrown-Norway Rats

Animal models of asthma can be used to demonstrate the effectiveness ofCGP on asthma. The Brown-Norway (BN) rat model of airwayhyperresponsiveness (AHR) has been shown to possess features similar tohuman allergic asthma, such as early- and late-phase reactions afterallergen challenge and the development of an IgE response to allergicsensitization (Haczku et al., Immunology. 85(4):598-603 (1995)). Such BNAHR rats can be produced by sensitization and exposure to ovalbumin(OVA) following a method similar to that described in Eur J. Pharmacol.7; 293(4):401-12 (1995) or Haczku et al. (1995), above. CGP can beadministered to the BN AHR rat prior to or after irritating the BN AHRrats with OVA. The airway responsiveness can be measured prior to orafter CGP administration.

Brown Norway rats (8-10 weeks old; weight: 150-300 g) are bred andmaintained by adhering to standards/guidelines established by theAssociation for Assessment and Accreditation of Laboratory Animal Care(“AAALAC”). These rats are sensitized by giving 4 mg of aluminumhydroxylate and 1 mg ovalbumin (OVA; Sigma, grade V) in one ml ofpyrogen-free saline by cervical subcutaneous injection forsensitization. Bordetella pertussis vaccine containing 3×10⁹ heatinactivated bacteria is used for intraperitoneal injection as anadjuvant. As a control, the same solution described above but withoutovalbumin is intraperitoneally injected, thus providing a negativecontrol. About 10-15 days after sensitization, airway irritation isinduced by challenging the rats with 5-10% aerosolized OVA for 5-10 minusing a nebulizer. Aerosol exposure can be accomplished by placing therats in a plexiglass chamber connected to a nebulizer, which generatesan aerosol mist pumped into the exposure chamber by the airflow suppliedby a small animal ventilator set at 60 strokes/min, with a pumpingvolume of 10 ml.

CGP can be administered to the rats by nasal inhalation (using anaerosol or a powder), or intubation (liquid, aerosol, respirator),either before or after the airway irritation is induced.

Airway responsiveness of the rats can be measured. Anaesthetized,tracheostomized and ventilated rats are monitored for their airflow,transpulmonary pressure and blood pressure. Lung resistance issimultaneously calculated using known method, such as a software program(LabView, National Instruments, Austin, Tex.). Bronchial Alveolar Lavage(BAL) fluid is collected. BAL is performed using 5 ml of phosphatebuffered saline (PBS; 137 mM NaCl, 10 mM sodium phosphate buffer pH 7.4,2.7 mM KCl), 4 times. The total number of cells in BAL fluid can bedetermined using an erythrocytometer and the percentage of eosinophilsin BAL fluid can be determined by differential cell counting.

An OVA challenge will elicit a statistically significant increase ininfiltration of inflammatory cells mainly consisting of eosinophils inthe airway. The ability of CGP to suppress the increase of eosinophilsin BAL fluid in response to the OVA challenge will be measured. Theeffectiveness of CGP can be studied at various dosages, in variousformulations, at various pH, etc., together with proper controlformulations.

EXAMPLE 2 Effectiveness of Calcium Glycerophosphate in Allergic RhinitisAnimal Models

Animal models of rhinitis can be used to demonstrate the effectivenessof CGP on rhinitis. Methods are known to produce animal models ofrhinitis. For example, a mouse model of allergic rhinitis can bedeveloped by local sensitization of mice with Dermatophagoidespteronyssinus (Kim et al., Otolaryngol Head Neck Surg. 2007 May;136(5):720-5); and a guinea pig model of rhinitis can be developed bybilateral intranasal sensitization of guinea pigs with the instillationof cedar pollen extracts (Mizutani et al., Jpn J Pharmacol. 2001 June;86(2):170-82).

CGP can be administered to the animal models of rhinitis via desirablemeans such as inhalation or spray. Symptoms of rhinitis, nasal mucosaeosinophilia, serum total IgE, cytokines, and eosinophilia in BAL fluidof the animal models are measured before or after CGP administration inorder to evaluate the effectiveness of CGP.

EXAMPLE 3 Applications of Calcium Glycerophosphate to a Human SubjectHaving a Respiratory Condition

Investigation of the effectiveness of a calcium glycerophosphate fortreating or preventing a disease, disorder and/or condition of therespiratory system has now been extended further to the human body.

CGP was incorporated into samples of a lavage or a spray product atvarying concentrations, along with different sweetening/moisturizingcomponents (sorbitol, glycerin) at varying levels (0.5% to 7.5% byweight), appropriate food/drug grade fungistats and bacteristats (methylparaben and benzalkonium chloride), and light flavorings of severaltypes (peppermint, spearmint and orange) in some iterations.

A first human subject for laboratory/home trials was a 77 year-old manin good general health. The subject has a life history of childhoodasthma, which is currently inactive except for rare mild adult episodes,all of which occurred several years prior, deviated septum blocking ofone nostril ˜60%. The subject has a history of childhood allergies toragweed, house dust, horse hair, etc., but these are inactive atpresent. The subject has experienced a decades-long post nasal drip(PND) as an adult, nasal obstruction (mucus, septum). The subject hasexperienced coughs that have been of weeks-long and, on occasion,months-long duration, following seasonal flu or colds. The subject hasexperienced chronic throat-clearing from PND and chronic cough even whennot post-respiratory illness, which he ascribes to bronchitis. Thisresults in occasional inconvenience when the subject has had to excusehimself from a group to clear out throat elsewhere, including spittingout or reflexive swallowing of the mucus which will on occasion roughenvoice. The subject breathed almost exclusively through mouth as a childbecause of nasal obstruction. The subject breathes through both nose andmouth as an adult, with breathing through nose occasionally mildlyproblematic, requiring active conscious engagement, although the subjectwas never “short of breath” in terms of lung capacity, etc.

Forced Nasal Lavage Experiment

The subject performed forced nasal lavage once daily for about twoweeks. The forced nasal lavage was conducted intranasally over skin andnormally in a shower. An aqueous solution or suspension of CGP atvarious concentration, such as about 7.5%, 3.75%, 2.5%, 2%, 1.5% byweight of dry CGP, was forced into one nostril and out of the othernostril using a closed plastic bottle with an internal stem feed. Thisprocedure was repeated for each side of the nostrils. Some product wasswallowed via nasal route and there was approx. 2 oz. fluid per nostril.

Within about 0-10 minutes after the lavage, the subject experiencedconsiderable drainage fore and aft the nasal passages, considerablethroat-clearing, and very wet nasal passages. The subject spit out someswallowing. Within about 10-20 minutes after the lavage, the subjectexperienced 90% decreased drainage, open and drying nasal passages, andeasier breathing. Within about 20-60 minutes after the lavage, thesubject felt that nasal passages remained open and drying, voice wasclearer and throat-clearing was about 90% diminished compared with priorstate. Within about 1 hr to 12/24 hr after the lavage, the subjectexperienced no drip, no cough, and easy open breathing. He could breathenaturally through his mouth and nose without active consciousengagement. His lower airway passages opened fully.

Spray/Mist Experiments

The subject applied CGP into his nostrils in a spray or a mist. A closedsmall plastic bottle with internal stem feed constructed to deliverfluid contents as mist was used. Typically, one application wasperformed in the morning. Each application consisted of two sprays pernostril, alternating sides, with full simultaneous deep breathinhalation or inspiration to draw the product up the nose and into theairway passages. The spray was also, on occasion, applied directlythrough the mouth to the throat with 1 or 2 sprays per application. Anaverage spray delivery contained about 200 mg of a formulation of CGPper squeeze. The formulation contained about 2% to about 3.75% by weightof dry CGP.

The subject experienced benefits from the nasal spray essentially thesame as those of lavage but without all the preliminary hydraulics andrecovery from same. CGP exerted its relieving effects almostinstantaneously upon application and the benefits lasted about four toeight hours post application. The spray/mist application was repeatedoccasionally in the afternoon or as needed. It was observed that asingle spray per nostril at bedtime appeared to produce better nightbreathing. The spray/mist application was much neater, easier tooperate, more portable, and more feasible for regular use.

After administration of the CGP to the respiratory system, either viaforced nasal lavage or spray/mist, the subject experienced about 95-98%reduction of post nasal drip, a daily nuisance to subject whetherupright and active or reclining, complete cessation of coughing, openbreathing through his nose without any, or at worst, with minimalrestriction.

In addition to the routine application of CGP to prevent or controlsymptoms associated with the respiratory system, the subject has alsoused CGP to relieve symptoms during an episode of a condition ofairway/upper respiratory tract. In one morning, during early producttrials, after the subject discontinued use of CGP for about 24 hours,the subject appeared to exhibit the following symptoms associated withairway/upper respiratory tract: nasal congestion, throat irritation withpost-nasal drip, hoarseness, and throat clearing, faint breathingsounds, and slightly short of breath. Although the symptoms did notamount to an asthma attack, the subject experienced significantdiscomfort. At about 10:45 A.M., the subject applied a double-dose ofNasoCell (1.75% CGP by weight) via nasal inhalation in each nostril. Atabout 11:30 AM, the subject's symptoms were much improved. The subjectthen self-administered another dose of NasoCell in each nostril. Atabout 12:50 P.M., it appeared that the subject's symptoms had almostcompletely resolved. The subject had no trouble breathing while he waseating during lunch. At about 2:30 PM, the subject's symptoms werecompletely resolved. The subject's throat was cleared of anyobstructions and breathing sounds, and was wide open. The subject'snasal passages were reopened and voice cleared from an earlier morninghuskiness, characteristic of a post nasal drip condition. It isnoteworthy that this subject observed that after some weeks of usage,much reduced usage was required to “maintain”, and that this includedthe skipping of usage altogether, some days.

EXAMPLE 4 Applications of Calcium Glycerophosphate to a Human SubjectHaving Asthma

This Example describes a study of the effectiveness of calciumglycerophosphate to treat or prevent asthma in a human subject. A CGPcomposition similar to that described in Example 3, containing about 2%(w/w) CGP, was used in this study. A second human subject, a man of 44years of age having a life history of asthma, participated in thisstudy.

The subject intentionally stopped using Advair Diskus (steroid) forabout 2 weeks. His allergies to molds were triggered by a ‘musty’ indoorenvironment resulting from several rainy and humid days. The allergiesin turn triggered his asthma. The subject had felt a slight reduction inhis airflow capacity for a few days, but had withheld use of albuterolinhaler (asthma rescue medication) during the day and only used it forsevere wake-ups during the night. The subject had to use albuterolinhaler for three contiguous nights prior to the night described below.

In that night, at about 8:30 PM, the subject was experiencing difficultybreathing and wheezing, which signaled the start of an asthma attack. Atabout 8:35 PM, the subject applied two sprays of the CGP compositioninto each nostril and one spray of the CGP composition into his mouthfor inhalation. An average spray delivery contained about 200 mg of thecomposition. At about 8:40 PM, the subject repeated the oral inhalationonce, and the subject experienced mild relief to the extent that he didnot feel the urgent need to use the albuterol inhaler. At about 8:52 PM,the subject did two additional oral inhalations of the composition. Atabout 8:55 PM, the subject noticed that his wheeze and tightness inchest were relieved about 80% or more. All nasal drips had been sniffedback in. At about 8:58 PM, the subject perceived a gradual improvementand experienced no labor in breathing. At about 9:05 PM, the subjectfelt better, but desired full relief and wanted to increase breathingcapacity. He applied one inhalation of the composition into eachnostril; paused to get full breath, then applied two deep oralinhalations of the composition at about 9:07 PM. At about 9:27 PM, thesubject felt fine and substantially back to normal. At about 9:37 PM,the subject felt improved. At about 10:16 PM, the subject felt fully(100%) recovered. At about 11:00 PM, the subject felt fine, no worry,and went to bed. The subject experienced no wake-up with shortness ofbreath and did not use inhaler during the night. At about 8:00 AM thenext day, the subject felt a little tightness in chest. Instead of using2 puffs of albuterol to start the day as he normally did, the subjectused the CGP composition. He applied two sprays of the composition intoeach nostril and two sprays of the composition into his mouth forinhalation. He felt relief and no need to use the albuterol inhaler.

EXAMPLE 5 Effectiveness of Calcium Glycerophosphate in Allergic RhinitisAnimal Models

This Example describes a study to determine whether repeated inhalationand intranasal instillation of calcium glycerophosphate attenuatesragweed induced nasal congestion and has an effect on nasalinflammation.

This study complied with all applicable sections of the Final Rules ofthe Animal Welfare Act regulations (9 CFR Parts 1, 2, and 3) and theGuide for the Care and Use of Laboratory Animals (National ResearchCouncil, 1996) with a study protocol reviewed and approved by theInstitutional Animal Care and Use Committee (IACUC) before theinitiation of the study. This study was conducted in the spirit of U.S.FDA 21 CFR Part 58 (Good Laboratory Practice for Nonclinical LaboratoryStudies), even though not all study aspects were within strictcompliance.

Beagle dogs immunized with ragweed (RW) as puppies develop allergicimmune responses. These allergic dogs show elevated total and specificserum IgE and increased numbers of eosinophils in their blood and lungs,as well as an increase in airway resistance and a decrease in dynamiccompliance after a challenge with RW by inhalation or local instillationin defined lung lobes. Additionally, these dogs develop increased nasalcongestion and inflammation following RW challenge in the nose.

Ragweed sensitized beagle dogs (n=5) with preexisting nasal and airwayallergic responses were utilized in the animal study described in thisExample. Dogs served as their own control and initially were treatedwith vehicle (sterile water) and then rested for 5 weeks prior toreceiving the test compound. The target dose of calcium glycerophosphatewas about 30 mg/dog twice daily. The maximum concentration of thecompound used for inhalation and nasal instillation was 20 mg/ml due tosolubility issues. Based on the aerosol testing only 1.6 mg/dog could bedelivered by inhalation during a 15 minute exposure (20 mg/ml solution).In addition, each dog received twice daily an intranasal instillation of30 mg (2×375 μl of 20 mg/ml in each nostril) given within 30 minutesafter the end of the inhalation exposures.

FIG. 1 summarizes the study design for characterizing the effects of CGPon rhinitis symptoms in ragweed sensitized Beagle dogs. The baselinenasal cavity volume was first measured 3 days (day −3) before each ofthe vehicle and CGP treatments. The dogs were treated with vehicle orCGP (day 0) twice daily with the target dosage described above. Threedays into the vehicle or CGP treatment, nasal challenge with ragweed wasperformed (day 3). Treatment with vehicle or CGP continued during theassessment of rhinitis endpoints, e.g., total treatment duration=6 days(day 0-5) for each of the vehicle and CGP treatments. There were 5 weeksfollowing the first intranasal RW challenge during the vehicle treatmentand the CGP treatment.

Vehicle (sterile water) was given twice daily by inhalation (15 minutes)followed by intranasal instillation of water (2×250 μl in each nostril).The test compound calcium glycerophosphate was formulated in sterilewater at a dose of 20 mg/ml and given by inhalation (deposition 1.6 mg).The test-article and the compound solution were prepared in the morningof each treatment day. Vehicle and test-article were administered byinhalation and intranasal instillation twice a day. Dosing of vehicle ortest compound was achieved by utilizing a face mask connected to a jetnebulizer for inhalation delivery followed by intranasal instillation inboth nostrils using an Accuspray device.

Treatment occurred on Days 0 to 5 (only AM on Day 5). Vehicle andcompound were delivered in the morning (8:15-9:10 am) and the afternoon(2:00-3:30 pm). On days when acoustic rhinometry procedures wereperformed (Days 0), the morning dosing occurred such that the procedureswere performed within 1-6 h after dosing. The pH of the compound (asadministered for nasal inhalation) was recorded prior to administration.As identified in the aerosol exposure the average pH of the nebulizersolution was 8.85.

Blood sample was collected by placing a venous (cephalic or saphenous)catheter in the leg prior to anesthesia or by venipuncture (jugular)using a syringe or needle-vacutainer. On the study Days −3 (baseline),3, and 5 two 2 to 3 ml blood samples were collected for vehicle andcompound experiment prior to exposures and intranasal instillation forchemistry and CBC analysis.

Rhinitis assessments were conducted using one or more of the followingtests.

-   -   Nasal cavity geometry (AcR) pre/post antigen challenge in both        nares        -   1. time 0 (just prior to RW instillation)        -   2. at 15, 30, 45, 60, 75, 90 min, 24 h and 48 h post RW            challenge    -   Nasal lavage of both nares at baseline and at 24 h (Day 4) and        48 h (Day 5) post RW instillation and only one side at 15, 30,        45 and 60 min post RW challenge (Day 3)        -   1. Release of histamine, leukotrienes and prostaglandins in            nasal lavage fluid baseline (prior to compound or RW            treatment/challenge), 15, 30, 45, 60 min and 24 h post RW            instillation        -   2. Nasal lavage cells from the baseline (Day−3), 24 h            (Day 4) and 48 h (Day 5) hour lavage were used for            determination of inflammatory cell differentials        -   3. Excess nasal lavage fluid was frozen for analysis of            cytokines.    -   Cardiovascular readouts (heart rate, O2 saturation, body        temperature) throughout nasal congestion measurement.

Measurement of Nasal Cavity Geometry by Acoustic Rhinometry—RhinitisAssessment

Acoustic rhinometery was measured on experimental Days 3, 4, and 5.Nasal cavity volume was measured in anesthetized dogs using anEccovision Acoustic Rhinometry System (Hood Laboratories, Inc.,Pembroke, Mass.). Briefly, a wave tube containing a spark soundgenerator was connected with the nasal cavity using a plastic nosepiece. Based on nasal cast impressions and X-ray measurements from thedog nasal cavity, a distance from the nostril opening into the nasalcavity of 10 cm was used for all experiments. Acoustic reflections wereconverted to area-distance function curves and used to determine nasalcavity volume. Heart rate and O₂ saturation were measured throughout theexperiment. Body temperature was checked occasionally. To avoid a rapiddrop in body temperature associated with general anesthesia dogs wereplaced on a water circulating heating pad during the experiment ifnecessary. Nasal cavity volume was measured before and at different timepoints after nasal RW challenge (both nares; 0, 15, 30, 45, 60, 75, and90 minutes, 24 h, 48 h post RW challenge). Nasal lavages were performedat specific time points before and after RW challenge in both nares atbaseline (Day −3), and at 24 (Day 4) and 48 h (Day 5) post RW challengeand only one side at 15, 30, 45, and 60 min post challenge (Day 3).

Anesthesia—Rhinitis Assessment

Dogs were anesthetized with isoflurane (5% induction; 1 to 1.5%maintenance). Briefly, custom made face masks constructed out of rubbermaterial were placed over the muzzles of the dogs for the induction ofanesthesia to avoid isoflurane entering the nasal passages. The facemasks had a hole cut in the end to allow a brass tubing to protrude outapproximately 1 to 2 cm while sealing around its outside. The face masksalso occluded the nares of the dogs, thus, assuring mouth-onlyinhalation of the anesthetic. After inducing anesthesia, an endotrachealtube was placed in the trachea and anesthesia was maintained withisoflurane throughout the experiment. Dogs were placed in a supineposition for the nasal congestion measurements and in a prone positionwith a slightly tilted head for the nasal lavage procedure.

Nasal Ragweed Challenge by Instillation

Ragweed extract (RW short, Ambrosia artemisifolia, Greer, Lenoir, N.C.;6 mg/ml in 0.25 ml PBS) was instilled in both nares using an Accuspraydevice (Becton Dickinson).

Nasal Lavage

While under anesthesia a flexible plastic catheter was inserted severalcentimeters into the dog's nare. The nare was washed with a phosphatebuffered saline solution (PBS; 3×5 ml for collection of cells [baseline,24 and 48 h post RW challenge]; 1×5 ml for collection of fluid formediators [15, 30, 45, and 60 minutes post RW challenge]).

Pathologic Analyses

Total nasal cells were determined using an automatic cell counter. Cellswere spun onto slides by cytocentrifugation and stained with a modifiedWright-Giemsa stain. At least four hundred inflammatory cells (or lessif applicable) were counted and the percentage of specific cell typesdetermined for each animal (slides in duplicates, 200 cells per slide).The first lavage fluid sample (after centrifugation) was frozenseparately for mediator analysis. Mediator analysis for histamine,leukotrienes, and prostaglandins were performed according to the kitmanufacturer instructions (Immunotech—Beckman Coulter Company #IM2015,Neogen Corporation #406410, Cayman Chemical Company: PGE2—#514010,PGD2—#512011, respectively).

Daily Observations

Animals were examined twice per day (morning and afternoon) on each dayof the study. Examination was oriented toward identifying the onset andprogression of any abnormal clinical signs. No adverse health effectswere found during the duration of the experiments.

Body Weights

All animals were weighed on study Days −3, 3 and 5 only for compoundexperiment (Table 1).

TABLE 1 Body weight (kg) for all dogs during vehicle and compoundtreatment. Vehicle Calcium Glycerophosphate Animal Day −3 Day 3 Day 5Day −3 Day 3 Day 5 1 10.5 10.8 NA 10.8 10.4 10.4 2 10.25 10.0 NA 10.710.6 10.55 3 12.5 12.7 NA 13.0 13.3 13.2 4 10.5 10.3 NA 10.4 10.3 10.4 510.0 9.9 NA 9.8 9.9 9.8

Statistical Analysis

Changes in nasal cavity volume or mediator levels were assessed by twoway analysis of variance (ANOVA) with Bonferroni post-test. All otherstatistical comparisons were made using ANOVA with Dunnett's multiplecomparison test or paired two tailed t-test if appropriate. A value ofp<0.05 was considered statistically significant.

Other parameters were also measured from the dogs at various time pointsand compared between the treatments with vehicle and CGP.

A total of 5 ragweed-sensitized dogs were used in this experiment. Alldogs received vehicle first and after a 5 week recovery period, theywere treated with calcium glycerophosphate by inhalation followed byintranasal instillation. Due to solubility of the compound only about1.6 mg were deposited in the lung by inhalation and a total of 30 mgwere intranasally instilled using a BD accuspray device twice a day fora total of 5 days.

The results for all variables (change in nasal cavity volume andmediators in nasal lavage fluid) are shown over time as a comparisonbetween treatment group and vehicle before and up to 24 or 48 hoursafter treatment. The area under the curve (AUC) for the change in nasalcavity volume from baseline (0 to 90 minutes post ragweed) and the AUCfor the change in mediator levels over time (0 to 60 minutes postragweed) were calculated for each treatment. An increase in AUC forcompound treatment compared to vehicle indicates an increase in nasalcavity volume and therefore a decrease in nasal congestion (FIG. 3). Incontrast, a decrease in AUC for the mediator levels (e.g. histamine,leukotrienes, prostaglandins) indicates an attenuation of thesemediators due to compound treatment (FIGS. 5, 7 and 9). An area underthe curve of 150 is the equivalent of a 100% increase in nasal cavityvolume and no change in AUC (same as vehicle) indicates no change innasal cavity volume (=0%). The AUC is always shown as an averageincluding standard error for each treatment group and as changes foreach individual dog between vehicle and compound treatment.

As shown in FIGS. 2 and 3, repeated treatment (BID) with calciumglycerophosphate by inhalation (1.6 mg deposition) and intranasalinstillation (30 mg) starting three days prior to nasal ragweedchallenge significantly increased nasal cavity volume and thereforeattenuated nasal congestion compared to vehicle treatment (Two WayAnova, p<0.0001). The attenuation was statistically significant at alltime points from 15 to 90 minutes post ragweed (p<0.05, Bonferroniposttest).

It has been shown previously that treatment with a-adrenergic agonist,pseudoephederine (PSE; 3 mg/kg) and histamine H1 antagonist,chlorpheniramine (10 mg/kg), in the same manner as that in the presentstudy can prevent the development of RW-induced nasal congestion(Rudolph et al., 2003, Am J Rhinol., 17(4):227-32). The response ofcalcium glycerophosphate is similar to that observed previously with PSEand chlorpheneramine with respect to the attenuation of nasal congestioninduced by intranasal ragweed challenge.

The histamine, leukotriene and prostaglandin D₂ and E₂ levels in nasallavage fluid were partially altered after treatment with the compoundcompared to vehicle treatment. In general, the levels of histamine,leukotriene and prostaglandin D₂ in nasal lavage fluid increased overtime compared to baseline levels after vehicle treatment. Statisticalsignificance was not reached between vehicle and compound treatment byan analysis under One Way Anova followed by Dunnett' s posttest.

The variable effects of compound treatment on individual dogs, e.g.,with some dogs showing a negligible effect and others showing a moredramatic effect, have also been observed with other compounds. Ingeneral, the dogs' response to RW challenge remains consistent followingrepeated RW challenges, and decreases in nasal congestion are due tocompound treatment, not due to variability in the response over time. Ahigh degree of variability in the peak levels of mediators, e.g.,leukotrienes, prostaglandins, and histamine, has also been generallyobserved following RW challenge. In addition, other compounds that haveresulted in reduced nasal congestion have also been associated with orwithout significant reductions in the mediators.

Interestingly, PGE₂ levels were elevated in the dogs prior to theinitiation of the calcium glycerophosphate treatment leg (e.g. T₀[baseline] levels). Levels of PGE₂ decreased following ragweed challengeduring the calcium glycerophosphate treatment (FIG. 10). Upon RWchallenge the PGE₂ levels returned (T30-60 min) to levels measuredfollowing the vehicle treatment leg and then were elevated at 24 h postRW challenge. This decrease in changes in PGE2 levels, with the graphsnormalized for both treated and untreated groups' baselines, achievesstatistical significance in favor of the treated group. PGE₂ has bothinflammatory and anti-inflammatory properties. PGE₂ promotesvasodilation by activating cAMP-coupled EP2 receptors on vascular smoothmuscle and increases vascular permeability indirectly by enhancing therelease of histamine and other mediators from tissue leukocytes such asmast cells. As inflammation progresses, PGE₂ synthesis by macrophages isenhanced due to increased expression of COX-2 and PGE-synthase. PGE₂inhibits leukocyte activation and promotes bronchodilation throughactivation of G_(s)-coupled EP2 and EP4 receptors (Tilley et al., 2001,J Clin invest. 108(1): 15-23). The elevated levels prior to calciumglycerophosphate may be the result of the initial RW challenge duringthe vehicle leg, despite the 5 weeks of rest in between. When PGE2levels are normalized to baseline levels, there was an overall effect oftreatment with calcium glycerophosphate, e.g. a greater reduction inPGE2 levels following RW challenge in conjunction with calciumglycerophosphate treatment, even though there was no significant effectat any specific time point (Two-way ANOVA with Bonferroni post test;FIG. 11). The overall greater reduction of PGE₂ following RW challengein conjunction with calcium glycerophosphate treatment suggests thatPGE₂ may be a potential underlying mechanism for the effects of calciumglycerophosphate.

The total collected cells include inflammatory cells and epithelialcells (FIG. 12). Epithelial cells and all inflammatory cells are shownas percentage of cells counted (FIGS. 13 to 17, panel A) and as numberof cells based on total number of cells collected (FIGS. 13 to 17, panelB). In this study the assessment of the number of inflammatory cells inthe nasal lavage showed that pretreatment with calcium glycerophosphatedid tend to reduce the lung inflammatory cells compared to vehicletreatment, although none of the endpoints reached statisticalsignificance under the current analysis. In general, RW challengeresulted in an increase in eosinophils on Day 1 and Day 2 postchallenge. Levels of macrophages following vehicle treatment were notdetectable and were at negligible levels following calciumglycerophosphate treatment.

Treatment with calcium glycerophosphate was well tolerated by theanimals and no visible adverse clinical signs were observed or anysignificant effects on blood chemistry parameters.

Overall, the response seen following calcium glycerophosphate treatmentis similar to the responses previously observed with an α-adrenergicagonist, pseudoephederine, a histamine H1 antagonist, chlorpheniramine,and montelukast in this model. Results of the animal study furtherdemonstrate that CGP is effective to relive nasal congestion, thususeful for treating or preventing a disease, disorder and/or conditionof a respiratory system related to an obstructive or a restrictivecondition of the respiratory airway, such as rhinitis, asthma, and otherTh2 inflammatory conditions.

It will be appreciated by those skilled in the art that changes could bemade to the embodiments described above without departing from the broadinventive concept thereof. It is understood, therefore, that thisinvention is not limited to the particular embodiments disclosed, but itis intended to cover modifications within the spirit and scope of thepresent invention as defined by the appended claims.

We claim:
 1. A method of treating rhinitis, allergic asthma or a chronicobstructive pulmonary disease (COPD) in a subject in need thereof, themethod comprising administering to the respiratory system of the subjectan effective amount of calcium glycerophosphate, wherein the calciumglycerophosphate is orally administered to the respiratory system of thesubject in a composition formulated for oral administration in a dosageof about 440 μg to about 88 mg of the, calcium glycerophosphate peradministration.
 2. The method of claim 1, wherein the calciumglycerophosphate is administered to the respiratory system of thesubject by a spray, a quick-dissolving tablet, an oral inhalationsolution or suspension, a syrup, a mechanized intermittent fluid pulser,an atomizer, a vaporizer, or an insufflator.